Formulations and compositions of poorly water soluble camptothecin derivatives

ABSTRACT

A- and/or B-ring substituted camptothecin derivatives, which are poorly water soluble (less than 5 micrograms per milliliter of water), are highly lipophilic camptothecin derivatives (HLCD) and are very active against a variety of human cancers. Because of their very poor water solubility, HLCD have not been used to treat human patients with cancer due to the inability to administer sufficient quantities of the HLCD dissolved in a pharmaceutical formulation. This invention overcomes these limitations by teaching novel pharmaceutically acceptable HLCD formulations for the direct administration of HLCD to human patients with cancer. The claimed invention also describes the methods to create solutions of HLCD and antitumor compositions of HLCD to allow the administration of HLCD in sufficient amounts to treat human patients with various types of cancer. This invention is also directed to injectable sterile solutions, antitumor compositions, solutions and suspensions comprising N-methyl-2-pyrrolidinone and a highly lipophilic camptothecin derivative.

This application is a division of application Ser. No. 08/461,385, filedJun. 5, 1995, now U.S. Pat. No. 5,726,181.

BACKGROUND OF THE INVENTION

1. Field of the Invention

During the past three decades it has been observed that camptothecin(CPT) and most of the highly lipophilic derivatives of camptothecin(HLCD) in their lactone form are poorly water soluble. For example, lessthan 5 micrograms of drug will dissolve in one milliliter of water toform a solution at a pH of 2 to 6. A range of pH from 2 to 6 maintainsthe dissolved camptothecin in the lactone form. Camptothecin and many ofits poorly water soluble derivatives are known potent anticancer drugs,however, their very poor water solubility has prevented their use in thetreatment of human cancer. The potency of these anticancer drugs wasdetermined by their ability to inhibit in vitro and in vivo tumor cellgrowth. This invention solves the poor solubility problems ofcamptothecins and its derivatives. Thus, the purpose of this inventionis to overcome the poor solubility of highly lipophilic camptothecinderivatives in their lactone form by designing novel formulations of thedrug (at sufficient concentrations) which can be administered orally,topically or parenterally for the purpose of treating human patientswith cancer.

2. Description of the Related Art

Introduction

A. DNA Topoisomerases

Several clinically important anticancer drugs kill tumor cells byaffecting DNA topoisomerases. Topoisomerases are essential nuclearenzymes that function in DNA replication and tertiary structuralmodifications, such as overwinding, underwinding, and catenation, whichnormally arise during cellular replication, transcription, and perhapsother DNA processes. Two major topoisomerases have been identified, bothof which are ubiquitous to eukaryotic cells: (1) Topoisomerase I (topoI) cleaves single stranded DNA; and (2) Topoisomerase II (topo II)cleaves double stranded DNA. Topoisomerase I is involved in DNAreplication; it relieves the torsional strain introduced ahead of themoving replication fork.

Topoisomerase I, purified from human colon carcinoma cells or calfthymus, has been shown to be inhibited by camptothecin and many of itsderivatives. Camptothecin, and water soluble camptothecin derivativesincluding CPT-11, topotecan, 9-amino camptothecin, 9-nitro camptothecin,DX8951 and 7-(4-methylpiperazinomethylene)-10,11-methylenedioxycamptothecin, 10,11-methylenedioxy camptothecin and 10,11-ethylenedioxycamptothecin have either been studied preclinically or used in clinicaltrials to treat certain types of human cancer. To date, there have beenno clinical studies in human patients involving poorly water solublehighly lipophilic camptothecins, other than for camptothecin (in thelate 1970's).

This absence of clinical use of lipophilic camptothecins has been due tothe lack of pharmaceutical formulations which allow the directadministration of the poorly water soluble camptothecin lactone speciesto human patients with cancer. For the purpose of this invention,examples of highly lipophilic camptothecin derivatives includecamptothecin, 10-hydroxy-7-ethyl camptothecin (SN38), 7-ethylcamptothecin (SN22), 10,11-methylenedioxy camptothecin,10,11-ethylenedioxy camptothecin and other poorly water solublederivatives of camptothecin which are active antitumor agents.

For the purpose of this invention, poorly water soluble and highlylipophilic camptothecin derivatives (referred to as "HLCD" for thepurposes of this invention) are defined interchangeably as any A- and/orB-ring substituted camptothecin which have a water solubility of lessthan 5 micrograms per milliliter of water. Also for the purposes of theinstant invention, the terms "highly lipophilic" and "poorly watersoluble" are used interchangeably to describe their fundamentalbioavailability and chemical behavior. Poorly water soluble camptothecinderivatives use the same mechanism to inhibit Topo I; they stabilize thecovalent complex of enzyme and strand-cleaved DNA, which is anintermediate in the catalytic mechanism. These compounds have no bindingaffinity for topoisomerase I but do bind with measurable affinity to theenzyme-DNA complex. The stabilization of the topoisomerase I "cleavablecomplex" by camptothecin and its derivatives is readily reversible.

Although camptothecin and the aforementioned poorly water solublecamptothecin derivatives have no effect on topoisomerase II, thesecamptothecin derivatives stabilize the Topoisomerase I--DNA "cleavablecomplex" in a manner analogous to the way in which epipodophyllotoxinglycosides and various anthracyclines inhibit topoisomerase II.

Inhibition of topoisomerase I by camptothecin and highly lipophiliccamptothecin derivatives induces protein-associated DNA single-strandbreaks. Virtually all of the DNA strand breaks observed in vitro cellstreated with camptothecin are protein linked. However, an increase inunexplained protein-free breaks can be detected in L1210 cells treatedwith camptothecin. The compounds appear to produce identical DNAcleavage patterns in end-labeled linear DNA. It has not beendemonstrated that camptothecin or highly lipophilic camptothecinderivatives cleaves DNA in the absence of the topoisomerase I enzyme.

B. Activity of Highly Lipophilic Camptothecin Derivatives is Cell CycleSpecific

The activity of highly lipophilic camptothecin derivatives is cell cyclespecific. The greatest quantitative biochemical effect observed in cellsexposed to camptothecin and its derivatives is DNA single-strand breaksthat occur during the S-phase. Because the S-phase is a relatively shortphase of the cell cycle, longer or repetitive exposures to the drugsresults in increased cell killing. Brief exposure of tumor cells to thedrugs produces little or no cell killing, and quiescent cells arerefractory. These results are likely due to two factors:

(1) The drugs inhibit topoisomerase I reversibly. Although they mayproduce potentially lethal modifications of the DNA structure during DNAreplication, the breaks may be repaired after washout of the drug; and

(2) Cells treated with topo I inhibitors, such as camptothecins tend tostay in G0 of the cell cycle until the drug is removed and the cleavedDNA is repaired. Inhibitors of these enzymes can affect many aspects ofcell metabolism including replication, transcription, recombination, andchromosomal segregation.

C. Lactone Form of Highly Lipophilic Camptothecin Derivatives IncreasesAntitumor Activity and Reduces Water Solubility

Utilizing HPLC and NMR techniques, researchers have demonstrated thatcamptothecin and many of it's derivatives undergo an alkaline,pH-dependent hydrolysis of the E-ring lactone. The slow reactionkinetics allows one to assess whether both the lactone and non-lactoneforms of the drug stabilizes the topoisomerase I-cleaved DNA complex.Studies indicate that only the closed lactone form of the drug helpsstabilize the cleavable complex. This observation provides reasoning forthe high degree of drug activity observed in solid tumor models. Tumorcells, particularly hypoxic cells prevalent in solid neoplasms, haverelatively lower intracellular pH levels than normal cells. At pH levelsbelow 7.0, the lactone E-ring form of camptothecins predominates. Thus,the inventors believe that camptothecins will be more effective atinhibiting topoisomerase I in an acidic environment than in cells havinghigher intracellular pH levels.

One of the objects of this invention is to provide lactone stable poorlywater soluble camptothecin derivatives as the basis of the claimedsubject matter. For this invention, lactone stable camptothecinderivatives are defined as poorly water soluble A- and/or B-ringsubstituted camptothecins which are dissolved inN-methyl-2-pyrrolidinone (referred to as "NMP") in the presence of anacid with or without additional excipients as desired. The inventorshave discovered that highly lipophilic camptothecins display anunusually high degree of solubility (greater than 1.0 mg permilliliter)in N-methyl-2-pyrrolidinone (referred to as "NMP"). NMP, as apharmaceutical excipient, is safe for human administration and has beenfound by the inventors to be chemically inert with respect to poorlywater soluble camptothecins. The presence of an acid in the solutionfurther stabilizes the lactone E-ring form of the HLCD; this isparticularly useful when additional excipients are used and when thedrug formulation is diluted with aqueous media. For the purpose of thisinvention, lactone stable camptothecin and highly lipophiliccamptothecin are used interchangeably.

D. Camptothecin and Highly Lipophilic Camptothecins

In 1966, Wall and Wani isolated camptothecin from the plant, Camptothecaacuminata. In the early 1970's camptothecin reached Phase I and Phase IIhuman trials and was found to have antitumor activity, but it causedunpredictable myelosuppression and hemorrhagic cystitis. It is importantto note that all of these studies used sodium hydroxide formulations ofcamptothecin which greatly increased the water solubility of themolecule due to base mediated hydrolysis of the lactone E-ring to formthe carboxylate species of camptothecin in appreciable quantities. Atthat time, however, it was not recognized that the lactone E-ringspecies of camptothecin had significantly (e.g., greater than 10 fold)greater anti-tumor activity than the carboxylate form of camptothecin.Phase II studies with sodium camptothecin were limited because patientsgiven sodium camptothecin experienced unpredictable and severemyelosuppression, gastrointestinal toxicity, hemorrhagic cystitis, andalopecia. Clinical trials with sodium camptothecin (referred to as"SCPT" for the purposes of this invention) were eventually discontinuedbecause of these unpredictable toxicities and the lack of consistentantitumor activity.

To demonstrate the utility and novelty of the present invention, it isuseful to review the literature on human clinical trials conducted withSCPT administered parenterally to human patients with cancer. Gottlieband coworkers (Cancer Chemotherapy Reports 54:461; 1970) reported onclinical studies with the sodium salt of camptothecin (SCPT) which werebegun at the Baltimore Cancer Research Center in January 1969. In thisclinical trial, SCPT was administered as a rapidly running i.v. solutionover a 5-10 minute period at a concentration of 2 mg of SCPT permilliliter of saline. Doses of SCPT ranged from 0.5 to 10.0 mg/kg ofactual or ideal body weight (whichever was less). These investigatorsreported that because hemorrhagic cystitis was noted in several of theearly trials, patients receiving camptothecin sodium were well hydratedeither i.v. or orally for 72 hours after drug administration. It isinteresting to note that the mean urine recovery of camptothecin was17.4% over the first 48 hours (range: 3.6-38.9%) with most of the drugexcretion occurring in the initial 12 hours. When these investigatorsexcluded the 5 patients with impaired urinary excretion, the mean urinerecovery of camptothecin was 22.8%. These investigators noted thatunmetabolized camptothecin in high concentrations rapidly appeared inthe urine after i.v. drug administration and went further to state thatthis finding probably accounted for the sterile hemorrhagic cystitisnoted in 3 moderately dehydrated patients. Although maintaining acopious urine outflow seemed to prevent this complication, theinvestigators explored various alterations in urine pH as anotherpossible way of decreasing the risk of this debilitating type oftoxicity.

Muggia et. al. (Cancer Chemotherapy Reports 56:515; 1972) reportedresults of a Phase I clinical trial in fifteen patients treated withSCPT at four weekly dose levels ranging from 20-67 mg/m². No clinicalbenefit was observed in eight patients with measurable disease who weretreated with the 5-day courses at dose levels associated with toxicity.The drug was administered in concentrations of 1 to 10 mg/ml and it wasalways administered by intravenous push. Cystitis was the most prominentnon-hematologic toxic effect observed in this study. Bladder toxicitywas dose limiting in three patients receiving doses of 20 to 30 mg/m²,and occurred in two additional patients at doses of 44 and 30 mg/m².Cystitis, another toxic effect occurring frequently after treatment withSCPT, was not predicted by preclinical toxicologic studies. Theirclinical experience suggested that the occurrence of cystitis may berelated to the duration of the patient's exposure to the drug whenadministered as the carboxylate form. Camptothecin is excreted unchangedby the kidneys, although a large percentage of the drug administeredcannot be accounted for in the urine and is likely conjugated in theliver to form the glucuronide and excreted via the hepatobiliary route.It is possible that relatively less drug is excreted in the urine ofanimals since an extremely active transport of camptothecin into bilehas been demonstrated. Alternatively, these investigators postulatedthat the mucosa of the human bladder is more susceptible to the toxicaction of camptothecin or that the effect on the human bladder is due tosome unrecognized camptothecin metabolite.

Moertel and coworkers reported results of a Phase II Study ofCamptothecin (NSC-100880) in the Treatment of Advanced GastrointestinalCancer (Cancer Chemotherapy Reports 56:95; 1972.) These investigatorsadministered camptothecin sodium dissolved in physiologic saline at aconcentration of 2 mg/ml and administered by rapid intravenous infusionover 5-10 minutes. Two schedules of administration were used in thisstudy: (a) a single injection repeated at 3-week intervals; and (b) a5-day course repeated ever 4 weeks. The initial dose for the single-dosemethod was 180 mg/m². Because of toxic effects which were consideredexcessive by the investigators, later patients were treated at dosesranging between 90 and 120 mg/m². Dosages for the 5-day course rangedbetween 11 and 22 mg/m² /day (total course, 55-110 mg/m²). Diarrhea wasonly a problem at higher doses, but then could be quite severe to thepoint of fecal incontinence and persistent for as long as 4 weeks.Cystitis usually began about 7-10 days after treatment and wascharacterized clinically by dysuria and frequency. With more severetoxicity, gross hematuria developed. Pathologically, this wascharacterized by multiple necrotic ulcerations which could involve theentire urinary tract from kidney pelvis to bladder. According to theseinvestigators, the occurrence of hemorrhagic cystitis did not precludefurther treatment with camptothecin, and its severity could be titrateddown by lowering the dose in subsequent courses. These investigatorsalso reported that the more prolonged schedule produced more severetoxicity at a given total dose level, but the difference was not asgreat as might have been predicted by preclinical animal studies. Theseinvestigators proposed that a reasonable initial dose of SCPT is 110-120mg/m² for the single-injection method or 17 mg/m² /day (total dose, 85mg/m²) for the 5-day course. They noted that after 2 months (8 or 9weeks) only two of their 61 patients showed evidence of partialobjective improvement, and none showed improvement at 3 months. Bothpatients who demonstrated an objective response at 2 months had largebowel cancer. These investigators concluded that camptothecin "is a drugof protean and unpredictable toxicity that has no clinical value in themanagement of gastrointestinal cancer." See Tables 1, 2, 3, and 4.

                  TABLE 1    ______________________________________    Toxic Reactions: Single-Dose Administration of    Sodium Camptothecin (Moertel et. al. Cancer Chemotherapy    Reports 56:95; 1972.)    Nonhematologic Toxicity No. of patients with:             No. of    Dose     patients    (mg/m.sup.2)             treated      Diarrhea Cystitis    ______________________________________    90       10           --       1    100      6            --       2    110      2            1        1    120      7            4        2    180      9            2        3    ______________________________________

                  TABLE 2    ______________________________________    Toxic Reactions: 5 Consecutive Day Administration    of Sodium Camptothecin (Moertel et. al. Cancer Chemotherapy    Reports 56:95; 1972.)    Nonhematologic No. of patients with:              No. of    Dose      patients    (mg/m.sup.2 × 5)              treated      Diarrhea Cystitis    ______________________________________    11        2            --       1    15        9            1        4    17        5            4        2    20        10           4        6    22        1            1        --    ______________________________________

                  TABLE 3    ______________________________________    Relationship of Method of Administration to Cystitis    (Moertel et. al. Cancer Chemotherapy Reports 56:95; 1972.)    Method of administration               Single dose 5-Day course    Cystitis   (% of 34 patients)                           (% of 27 patients)    ______________________________________               24          48                           (P < 0.05)    ______________________________________

                  TABLE 4    ______________________________________    Objective Responses (Moertel et. al. Cancer    Chemotherapy Reports 56:95; 1972.)    ______________________________________    Single-dose method (34 patients)                 Time after start of therapy    Objective Responses*                   3 wks  6 wks    9 wks                                        12 wks    ______________________________________    Improved       4      2        2    --    Stable         17     11       8    6    Worse          13     21       24   28    ______________________________________    5-Day course (27 patients)                Time after start of therapy    Objective Responses*                  4 wks      8 wks  12 wks    ______________________________________    Improved      1          --     --    Stable        12         7      6    Worse         14         20     21    ______________________________________     *3 patients showed 25%-50% response at 3 wks only.

Gottlieb and Luce reported the results of treatment of patients withmalignant melanoma with camptothecin sodium (Cancer Chemotherapy Reports56:103,1972). Fifteen patients with advanced malignant melanoma weretreated with SCPT at doses of 90-360 mg/m² repeated every 2 weeks. SCPTwas administered as a single rapid intravenous (i.v.) injection startingat a dose of 120 mg/m² repeated at 2-week intervals. The dose insubsequent courses was increased by increments of 60 mg/m² per dose (toa maximum of 360 mg/m²) in eight patients who tolerated their initialdoses with minimal toxicity. To prevent the known bladder toxicity ofthis drug, patients were well hydrated for 3 days after therapy. None ofthe patients had a 50% or greater decrease in tumor diameter. Lesspronounced transient tumor regression was noted in three patients, butno clinical benefit was associated with these responses. The remainingpatients had no change or progression in their disease. Toxic effectsincluded myelosuppression (11 patients), nausea and vomiting (9patients), alopecia (8 patients), diarrhea (3 patients), and hemorrhagiccystitis (1 patient). These investigators concluded that SCPT, at leastas administered in this study, had little to offer the patient withadvanced disseminated melanoma.

Creaven and co-investigators reported studies of plasma camptothecinlevels during a 5 consecutive day course of treatment (CancerChemotherapy Reports 56:573-578, 1972). These investigators state thatthe toxicity of SCPT has been widely and unpredictably variable in thecourse of initial clinical evaluation. Severe toxic effects includingcystitis occurred even though patients with obvious renal disease wereexcluded. In this study they investigated plasma camptothecin levels 24hours after the administration of SCPT administered on a once daily×5schedule to determine whether such measurements would be of value inpredicting toxicity, and observed that plasma camptothecin levels havelittle relation to the dose given when the dose is in the range of6.5-20 mg/m² /day.

In another clinical study Muggia and co-workers reported the results ofa Phase I Trial of weekly and daily treatment with SCPT (CancerChemotherapy Reports 56: 515-521, 1972). Fifteen patients were treatedat four weekly dose levels ranging from 20 to 67 mg/m² of SCPT.Reversible leukopenia was the major dose-limiting toxic effect. Five-dayloading courses were begun at total doses of 1.5 mg/m² per coursebecause increased sensitivity to daily administration had been noted inanimal studies. Leukopenia was more prolonged after daily treatment thanafter weekly treatment and occurred in four of six patients receiving atotal dose of 100 mg/m². Tolerance to 5-day courses was an unexpectedclinical result. Also unpredicted by preclinical studies was humansusceptibility to cystitis with either schedule of treatment. They notedclinical responses in two of ten patients in whom responses could beevaluated after weekly courses of treatment. No clinical benefit wasobserved in eight patients with measurable disease who were treated withthe 5-day courses at dose levels associated with toxicity. Cystitis wasanother toxic effect occurring frequently after treatment with SCPT, andthis toxicity was not predicted by preclinical toxicologic studies. Theinvestigators suggested that the occurrence of cystitis may be relatedto the duration of the patient's exposure to the drug, and proposed thatcamptothecin is excreted unchanged by the kidneys, although a largepercentage of the drug administered cannot be accounted for in theurine. They also proposed from this study that it is possible thatrelatively less drug is excreted in the urine of animals since anextremely active transport of camptothecin into bile had beendemonstrated. They also postulated that the mucosa of the human bladderis more susceptible to the toxic action of camptothecin or that theeffect on the human bladder is due to some unrecognized camptothecinmetabolite.

There are several features which are common in these earlier clinicalstudies with SCPT. First is the use of SCPT ("SCPT") which made thecamptothecin more water soluble. Hydrolysis of the lactone E-ring toform the water soluble carboxylate species was accomplished byformulating camptothecin in sodium hydroxide. The antitumor activity ofthe carboxylate form of camptothecin is reduced by at least 10-fold,which partially accounts for the lack of clinical response in thesestudies. Second is the rapid intravenous administration of the drug.Camptothecin is an S-phase specific drug and therefore will exert agreater antitumor effect under conditions of prolonged or repetitiveexposure, as in a continuous intravenous infusion or repetitive dailydosing. The short infusion (i.v. push or rapid i.v. infusion) times inall of these earlier studies do not allow a long enough exposure time tothe drug to attain suitable plasma drug levels, and is furthercompounded by the administration of the water soluble carboxylate formof camptothecin. A third common feature is the notable frequency ofcystitis in these studies using water soluble SCPT.

The novel features of the present invention includes the following: (1)pharmaceutically acceptable formulations which allow direct parenteraladministration of lactone stable highly lipophilic camptothecinderivatives to human patients with cancer (referred to as "HLCD"); (2)pharmaceutically acceptable formulations which allow the direct oraladministration of lactone stable HLCD to human patients with cancer. Theinventors predict that by administering the carboxylate species of HLCDa higher incidence of renal toxicity is likely to be observed than ifthe lactone species of HLCD is administered to patients.

The inventors maintain that the previous use of SCPT caused hemorrhagiccystitis relates to the enhanced renal excretion of the carboxylate formof camptothecin which when exposed to the lower pH (˜pH 6 or less) ofthe distal convoluted tubule and collecting duct in the kidney, assignificant proportions of the carboxylate form of camptothecin isconverted into the lactone form. The formation of the lactone species inhigh concentration at the distal convoluted tubule and collecting ductresulted in a high concentration of the lactone form of camptothecinbeing excreted and damaging the uroepithelium which resulted inhemorrhagic cystitis. Elimination of a greater concentration of thelactone form of camptothecin by the renal route is enhanced byadministration of the water soluble carboxylate form and is greatlyreduced by administration of the lactone form of the drug. Thus,additional significant utilities of the present invention are that theadministration of HLCD substantially in the lactone form orally orparenterally to cancer patients will significantly reduce the renalelimination of HLCD and further that the incidence of hemorrhagiccystitis will be significantly reduced in patients who receive theformulations of HLCD claimed in this invention.

In addition to the previously noted toxicities and limited clinicalresponses to camptothecin, HLCD have also been considered unsuitable forclinical use because they are all poorly soluble in water (e.g. lessthan 5 micrograms of HLCD per milliliter of water). The poor watersolubility of HLCD requires the use of large volumes of water-basedparenteral vehicle, which results in administering the drug for aprolonged period of time and causes inconvenience and discomfort topatients. Also, administering the drug for prolonged period of timeincreases the costs associated with treatment of patients. Hence, anHLCD formulation which permits higher concentrations of the active drugin the infusion after dilution with a suitable parenteral vehicle isdesired. Also desired is that the drug remains in the diluted solutionfor sufficient amount of time to be effective. Such an infusion solutionwill be greatly beneficial to patients, by bringing down the timerequired for administering useful amounts of drug and also the costsassociated with administering the drug for a more prolonged period oftime.

One highly useful purpose of this invention is to formulate the HLCD ina pharmaceutically acceptable manner using an organic or a mixture oforganic co-solvents with a high degree of physiologic safety to dissolvethe HLCD in desirable concentrations and concurrently stabilize HLCD inthe lactone E-ring form. It is this formulation invention which permitsdirect administration of HLCD to human patients with cancer.

The inventors believe that direct administration of a lactone stableHLCD to human patients has another important utility for treatingpatients with cancer. It is well known that cell membranes are comprisedlargely of lipid, and that lipid soluble drugs in general have superiorability to penetrate hydrophobic cellular membranes relative to watersoluble drugs. HLCD are poorly water soluble and are therefore lipidsoluble which facilitates their penetration into various body tissuesand will improve the bioavailability and anticancer activity of thedrug. The anticancer activity of the camptothecins in general is closelylinked with their ability to inhibit the intracellular Topoisomerase Iwhich is concentrated within the nucleus of the cell. The inventorscontend that the present invention increases the amount of HLCD drugdiffusion through the cellular and nuclear membranes in tumor cells andwill result in superior antitumor activity of the drug. Water solublecamptothecin derivatives are predicted to have a lesser ability todiffuse through the cellular and nuclear membranes in the body.

The utility of suitable organic solvents for this invention involvingpharmaceutical dosage forms of HLCD is restricted to those which have ahigh degree of physiological safety in humans. This invention teachesnew methods for making pharmaceutical formulations for a variety oflactone stable HLCD with water solubility of 5 micrograms per milliliteror less. Some examples of poorly water soluble camptothecins (HLCD)include 10,11-methylenedioxy camptothecin, 10,11-ethylenedioxycamptothecin, 7-ethyl camptothecin(SN22), 7-ethyl-10-hydroxycamptothecin (SN38) and congeners thereof. Any poorly water solublecamptothecin with a solubility of 5 micrograms per milliliter of wateror less may be dissolved or suspended in these novel formulations andwill have appreciable quantities (greater than 90%) of the lactone formof drug in the resulting solution. 10,11-Methylenedioxy camptothecin,10,11-ethylenedioxy-camptothecin, 7-ethyl camptothecin, and7-ethyl-10-hydroxy camptothecin and congeners thereof are reportedlyvery active in preclinical studies, but they are also reported to bepoorly soluble in water (less than 5 micrograms of drug will dissolve inone milliliter of water) which limits their utility because of theinability to readily administer these drugs to human patients withcancer (Pommier, et al. 1992, Wall et. al. 1994).

One of the advantages of the instant invention is that the instantformulations provide clinicians with the ability to directly adjust theplasma levels of HLCD to the point of therapeutic tolerance bycontrolling the dose and the schedule of drug administration. Theinventors contend that this should lead to a superior ability to achievemore effective antitumor activity and reduced interpatient variabilityof the plasma levels of HLCD.

The different observations made in these studies suggest that directadministration of HLCD by parenteral and oral administration couldprovide significant clinical benefit for patients undergoing treatmentfor cancer. However, in the past, HLCD have been consideredinsufficiently water soluble for clinical use. The current inventionovercomes the solubility problem by providing lactone stablepharmaceutically acceptable formulations of HLCD which upon dilutionwith an acceptable parenteral vehicle gives a stable solution of usefulconcentrations of HLCD for parenteral use and also a concentratedsolution or suspension of HLCD suitable for encapsulation within agelatin capsule for oral HLCD formulations.

SUMMARY OF THE INVENTION

This invention involves the pharmaceutical formulation of lactone stablehighly lipophilic camptothecin derivatives ("HLCD") to treat cancer inhumans. Also within the scope of the present invention is a stable HLCDsolution or suspension in NMP (1-Methyl-2-Pyrrolidinone, defined above)which, upon dilution with suitable parenteral vehicle, provides a finalinfusion containing a HLCD activity in the range of about 0.001 mg toabout 1.0 mg per ml. The present invention also relates to a highlyconcentrated solution or suspension of HLCD in the range of about 1.0 mgto about 40.0 mg per ml in NMP suitable for encapsulation within agelatin capsule. For the purposes of this invention, lactone stable HLCDis defined as any A- and/or B-ring substituted camptothecin derivative(HLCD) having a water solubility of less than 5 micrograms permilliliter of water in the lactone form.

For the purpose of this invention, a highly lipophilic camptothecinderivative ("HLCD"), having a water solubility of 5 micrograms permilliliter or less, has the general structural formula: ##STR1## whereinR₁, R₂, R₃, R₄ =H, lower alkyl, alkoxy, acyloxy, hydroxy, acyl, halo,amido, or cyano group;

wherein R₁ and R₂ together may represent --X₁ --X₂ --X₃ -- and whereinX₁, X₂, X₃ may be CR₅ R₆, O, S or NR₇ ; and wherein R₅, R₆, R₇ =H, loweralkyl, alkoxy, acyloxy, hydroxy, acyl, halo, amido, or cyano group;

wherein R₂ and R₃ together may represent --X₁ --X₂ --X₃ -- and whereinX₁, X₂, X₃ may be CR₅ R₆, O, S or NR₇ ; and wherein R₅, R₆, R₇ =H, loweralkyl, alkoxy, acyloxy, hydroxy, acyl, halo, amido, or cyano group;

wherein R₃ and R₄ together may represent --X₁ --X₂ --X₃ -- and whereinX₁, X₂, X₃ may be CR₅ R₆, O, S or NR₇ ; and wherein R₅, R₆, R₇ =H, loweralkyl, alkoxy, acyloxy, hydroxy, acyl, halo, amido, cyano group;

Another embodiment of this invention is an injectable, sterile solutioncomprising N-methyl-2-pyrrolidinone and a highly lipophilic camptothecinderivative ("HLCD") having a water solubility of 5 micrograms permilliliter or less, with the general structural formula: ##STR2##wherein R₁, R₂, R₃, R₄ =H, lower alkyl, alkoxy, acyloxy, hydroxy, acyl,halo, amido, or cyano group;

wherein R₁ and R₂ together may represent --X₁ --X₂ --X₃ -- and whereinX₁, X₂, X₃ may be CR₅ R₆, O, S or NR₇ and wherein R₅, R₆, or R₇ =H,lower alkyl, alkoxy, acyloxy, hydroxy, acyl, halo, amido, or cyanogroup;

wherein R₂ and R₃ together may represent --X₁ --X₂ --X₃ -- and whereinX₁, X₂, X₃ may be CR₅ R₆, O, or NR₇ and wherein R₅, R₆, or R₇ =H, loweralkyl, alkoxy, acyloxy, hydroxy, acyl, halo, amido, or cyano group; and

wherein R₃ and R₄ together may represent --X₁ --X₂ --X₃ -- and whereinX₁, X₂, X₃ may be CR₅ R₆, O, S or NR₇ and wherein R₅, R₆, or R₇ =H,lower alkyl, alkoxy, acyloxy, hydroxy, acyl, halo, amido, or cyanogroup.

A further embodiment of this invention is a pharmaceutically acceptableacid selected from the group consisting of acetic acid, citric acid,fumaric acid, maleic acid, ascorbic acid, hydrochloric acid, phosphoricacid, gluconic acid, lactic acid, and hydrochloric acid may also beadded to the above defined injectable sterile solution. For the purposesof this invention, a pharmaceutically acceptable acid is defined as anacid selected from the group consisting of, but not limited to, aceticacid, citric acid, fumaric acid, maleic acid, ascorbic acid,hydrochloric acid, phosphoric acid, gluconic acid, lactic acid, andhydrochloric acid.

Additionally, the above injectable sterile solution may also include oneor more excipients including, for example, but not limiting to, ethanol,benzyl alcohol, glycerin, polaxomer, PEG-300, PEG-400, Tween-80,Cremaphor or taurocholic acid or a pharmaceutically acceptable saltthereof.

For the purpose of this invention, one of ordinary skill in this artwould know that the A-ring is the first ring on the left side of theabove chemical structure and that the B-ring is the second from the leftring of the above structure. Additionally, one of ordinary skill in thisart knows that R-1 in the above chemical structure is also defined asposition 7 of the B-ring. R-2 in the above chemical structure is alsodefined as position 9 of the A-ring. R-3 in the above chemical structureis also defined as position 10 of the A-ring and R-4 in the abovechemical structure is also defined as position 11 of the A-ring.

Another embodiment of this invention is substitutions in only the A-ringof the above structure only, substitutions in only the B-ring only andalso substitutions in both the A-ring and the B-ring. Examples of HLCDA-ring substituted camptothecin derivative include, but are not limitedto, substitutions at positions 9, 10, or 11 and combinations thereof.For example, A-ring substitutions could be at position 9 only, atposition 10 only, at position 11 only, as well as A-ring substitutionsat positions 9 and 10, at positions 9 and 11, and at positions 9, 10,and 11. Additionally, B-ring substituted camptothecin derivativesinclude, but are not limited to, substitutions at position 7. Thisinvention also embodies substitutions in both the A-ring and in theB-ring. For example, this invention includes substitutions at positions7 and 9, at positions 7 and 10, at positions 7 and 11, at positions 7,9, 10, and 11 and at positions 7, 10, and 11. The above substitutionsare examples only and do not intend to limit the instant invention tothe substitutions listed.

Examples of HLCD (any A- and/or B-ring substituted camptothecinderivative) as defined in the instant invention include, withoutrestriction or limitation, 10,11-methylenedioxy camptothecin,10,11-ethylenedioxy camptothecin, 7-ethyl camptothecin,7-ethyl-10-hydroxy camptothecin, 9-methyl camptothecin,9-chloro-10,11-methylenedioxy camptothecin, 9-chloro camptothecin,10-hydroxy camptothecin, 9,10-dichloro camptothecin, 10-bromocamptothecin, 10-chloro camptothecin, 9-fluoro camptothecin, 10-methylcamptothecin, 10-fluoro camptothecin, 9-methoxy camptothecin, and11-fluoro camptothecin.

Direct administration of HLCD to human patients with cancer is likely tooffer several important clinical advantages over administration of morewater soluble camptothecin derivatives such as SCPT, CPT-11, topotecan,9-amino camptothecin, 9-nitro camptothecin and7-(4-methylpiperazinomethylene)-10,11-ethylenedioxy camptothecin. Forexample:

(1) direct administration of HLCD allows the clinician to tailor theadministration of the active cytoxic species (lactone stable form ofHLCD) to suit the patient's tolerance;

(2) direct administration of HLCD overcomes interpatient variabilitywhich may be due to polymorphism of key enzyme(s) in the metabolism ofCPT-11 to 7-ethyl-10-hydroxy camptothecin;

(3) clinicians can more consistently optimize the drug dosage andschedule to achieve the maximum tolerated dose of HLCD which is likelyto lead to the most beneficial clinical anti-cancer effect; and

(4) direct administration of an HLCD in the lactone form will have agenerally superior ability to penetrate tissue than the directadministration of water soluble camptothecin derivatives in the lactonestable or carboxylate forms.

Regarding the clinical utility of lactone stable pharmaceuticalformulations of HLCD for the treatment of human cancer, this inventionprovides the following:

(1) solutions and suspensions comprising lactone stable HLCD;

(2) formulations of lactone stable HLCD suitable for parenteraladministration;

(3) oral formulations of lactone stable HLCD; and

(4) use of formulations of HLCD for the treatment of localizedcomplications of cancer by direct administration via instillation intovarious body cavities.

HLCD Dissolved or Suspended in N-Methyl-2-Pyrrolidinone With or WithoutAdditional Excipients

Another embodiment of the claimed invention is a highly lipophiliccamptothecin derivative (HLCD) solution or suspension containing HLCDand N-methyl-2-pyrrolidinone ("NMP"). Yet another embodiment of theclaimed invention is a highly lipophilic camptothecin derivative (HLCD)solution or suspension containing HLCD, N-methyl-2-pyrrolidinone ("NMP")and a pharmaceutically acceptable acid.

There are many pharmaceutically acceptable acids for this invention, butthe inventors prefer to select one from the group consisting of aceticacid, citric acid, fumaric acid, maleic acid, ascorbic acid,hydrochloric acid, phosphoric acid, gluconic acid, lactic acid, andhydrochloric acid.

Taurocholic acid, a bile acid with very weak acidic properties may beused for certain oral formulations if desired, and is not incorporatedin any formulation for the purpose of lowering the pH of theformulation. Also for the purposes of this invention, the term"pharmaceutically acceptable" is defined as a reference to the highdegree of physiologic safety of the liquid organic excipients containedwithin the undiluted formulations when administered to human patients inthe amounts contained within a range of 1 to 50 milliliter volumesadministered to humans patients for one to five consecutive days.

One of the key discoveries in the present invention is the unexpectedlyhigh solubility of HLCD in N-methyl-2-pyrrolidinone (NMP).N-Methyl-2-pyrrolidinone is an organic liquid excipient and is alsoknown as 1-methylpyrrolidinone, N-methyl-2-pyrrolidinone,1-methyl-5-pyrrolidinone, methylpyrrolidinone, N-methyl pyrrolidinone,methylpyrrolidinone, N-methylpyrrolidone, N-methyl-2-pyrrolidone,M-pyrol, and NMP.

NMP exhibits a high degree of physiologic safety in mammals with thefollowing LD50 values: (rat) oral--7000 mg/kg, intraperitoneal--2472mg/kg, intravenous--2266 mg/kg, (mice) oral--7725 mg/kg,intraperitoneal--4429 mg/kg, intravenous--3605 mg/kg, (rabbit)skin--8000 mg/kg (Registry of Toxic Effects of Chemical Substances,1983-84 Supplement, Page 1628). NMP has been used to formulate etoposide(Etoposide, U.S. Pat. No. 4,772,589) and acridine derivatives (M-AMSA,U.S. Pat. No. 5,034,397). Etoposide and acridine derivatives (1) areanticancer drugs; (2) are chemically unrelated to HLCD; (3) are morewater soluble than HLCD; and (4) exert their antitumor effects by vastlydifferent mechanisms than HLCD.

NMP has also been used for oral formulations of the antibioticclarithromycin (Clarithromycin, WO patent #9,014,094) and other drugs.NMP is a key excipient of the instant invention which allows anexceptionally high degree. of drug solubility of HLCD (range 1 mg/ml to40 mg/ml)in NMP as a solution or suspension. An HLCD solution comprisingNMP with or without other combinations of excipients described herein,which can be diluted with a parenteral vehicle such as sterileinjectable water USP, 5% Dextrose solution for injection USP or 0.9%sodium chloride solution for injection USP, such that the amount of HLCDdissolved in the diluted infusion is from about 0.001 mg/ml to about 1.0mg/ml, is taught in the present invention. The inventors have discoveredthat HLCD show remarkably high solubility in NMP compared to othercommon pharmaceutical solvents such as water, ethanol, benzyl alcohol,propylene glycol, PEG 300, PEG 400, dimethylisosorbide ordimethylacetamide (Table 5). This high solubility of HLCD in NMP makesNMP a unique and highly useful pharmaceutical solvent for making usefulsolutions or suspensions of HLCD.

                  TABLE 5    ______________________________________    Solubility of Camptothecin in Various Solvents    Solvent             Concentration, mg/ml    ______________________________________    Milli-Q Water       0.002    Ethanol             0.051    Benzyl alcohol      1.674    Propylene glycol    0.281    PEG 300             0.706    Dimethylisosorbide  0.928    Dimethylacetamide   5.000    N-Methyl-2-pyrrolidinone (NMP)                        >15.000 (range 15-20)    ______________________________________

NMP is inert with respect to undesirable chemical reactions with HLCDand is therefore a highly useful excipient to create solutions of HLCDin the lactone form. Further utility of NMP in the present invention isthe discovery that NMP allows the introduction of additional excipientswhich further improve the overall utility of the HLCD dissolved in theNMP solution in a manner which are of additional benefit for parenteralor oral administration to human patients with cancer. The HLCD solutionor suspension is prepared by mixing the desired components with NMP andadding a pharmaceutically acceptable acid to adjust the pH to 3-5.

A pharmaceutically acceptable acid is preferably included in the NMPsolutions and suspensions of the present invention. Any pharmaceuticallyacceptable acid may be used; for example mineral acids such ashydrochloric acid or phosphoric acid; and carboxylic acids such astartaric, lactic, ascorbic, gluconic, citric, succinic, fumaric, ormaleic acids. Hydrochloric acid, phosphoric acid and carboxylic acidsare the most preferred for the novel oral and parenteral formulationsdescribed in the present invention. The amount of acid used may be fromabout 100 to about 5000 parts by weight of acid per part by weight ofHLCD and preferably from about 1000 to 2500 parts by weight of acid perpart by weight of HLCD. Citric acid is preferably used in a proportionof from about 1000 to about 2000 parts by weight. Oral formulations ofNMP and HLCD can additionally contain taurocholic acid. The NMP HLCDsolution is miscible with ethanol, benzyl alcohol, polysorbate-80(Tween-80), polyethylene glycol (PEG), polyoxyethylated castor oil,propylene glycol, isopropyl myristate, corn oil, cottonseed oil, and thelike.

An object of the present invention is to provide a solution orsuspension of HLCD in the lactone form in NMP. A more concentratedHLCD-NMP solution or suspension (10 mg or more per milliliter ofsolution or suspension) is particularly useful as a filling solution orsuspension for gelatin capsules. A HLCD-NMP solution may also beformulated for parenteral administration providing a useful andpractical means to dissolve the drug.

The present invention is prepared by mixing HLCD in NMP alone or bysubsequent addition of additional excipients including or excluding anycombination the following: (a) a carboxylic acid and/or mineral acid,(b) polyethylene glycol (PEG-300 and/or PEG-400), (c) alcohol (ethyland/or benzyl alcohol) (d) polysorbate-80 (Tween-80) and (e) taurocholicacid. The amount of HLCD contained in the solution or suspensiondescribed in this invention is not specifically restricted but may beany amount convenient for pharmaceutical purposes, and may be selectedaccording to the dosage to be prepared. A preferred capsule fillingsolution or suspension contains from about 1 mg to about 40 mg of HLCDactivity per ml of solution or suspension.

Another preferred embodiment of the claimed invention is an HLCDsolution or suspension prepared by dissolving or suspending the desiredcomponents in NMP in the presence of a pharmaceutically acceptable acid.

In the formulations provided by the instant invention, the HLCD issoluble or suspended and maintained in its active lactone form. Thenon-enzymatic conversion of the pH labile E-ring from the closed lactone(active) to the open carboxylate form (inactive) is reduced byformulating HLCD under acidic pH conditions (pH range of 3 to 5). Thus,an acid is included by the inventors to assure that an acidic pH valueis maintained upon dilution to form a micellar solution or suspension.Examples of preferred carboxylic acids effective in this inventioninclude citric, gluconic, lactic, maleic, tartaric, or ascorbic acids.Other acids such as hydrochloric acid and phosphoric acid can beemployed instead or in addition to citric acid to form the mostpreferred solution.

Yet another embodiment of the claimed invention is that the solution orsuspension of HLCD contains from about 1.0 mg to about 40.0 mg activityof HLCD per ml of solution or suspension. This concentration of HLCD inthe resulting formulation solution or suspension would be useful andeffective for both oral and parenteral administration of the HLCD tohuman patients with cancer.

When oral dosages are to be administered in a capsule form, it isadvantageous to have a concentrated solution or suspension of HLCDsuitable for encapsulation within a soft or hard gelatin capsule.Concentrated solutions or suspensions allow the preparation of capsulesof smaller size which allows easier ingestion by the patient, and mayalso reduce the number of capsules to be swallowed. These factors areimportant in view of the generally poor condition of cancer patients.

Taurocholic acid, a bile acid, may enhance in the intestinal absorptionof the drug in certain patients. The present invention takes advantageof the discovery that taurocholic acid, or a pharmaceutically acceptablesalt thereof, when included with HLCD in a solution or suspension dosagecomposition, results in improved absorption of the drug following oralingestion of the composition. It is believed that this is due to theformation of a micellar solution of HLCD on dilution thereof with thegastric contents.

The phenomenon of micellar solubilization of poorly water-soluble drugsmediated by bile acids, including taurocholic acid, has been previouslyreported with respect to glutethimide, hexesterol, griseofulvin (Bateset al.), reserpine (Malone et al.) and fatty acids and cholesterol(Westergaard et al.). The use of taurocholic acid or a pharmaceuticallyacceptable salt thereof in the present invention involves apharmaceutical solution of HLCD which has the useful property ofproviding a stable apparent solution of the drug upon dilution thereofwith from 1 to 100 volumes of water. The solution is stable and free ofprecipitate for a period of at least two hours; sufficient time topermit administration and absorption by the patient.

It has been observed with similar solutions of etoposide, which is achemically different anticancer drug, that the bioavailability of thedrug following oral administration is substantially equivalent to thatachieved by intravenous administration of a solution of the chemicallyunrelated anticancer drug etoposide (U.S. Pat. No. 4,713,246). Analogousto that of etoposide, it is believed that ingestion of the presentdosage form of HLCD and resulting dilution thereof by the stomachcontents, results in the formation of a micellar solution of HLCD in thestomach which is readily absorbed by the gastrointestinal tract.However, the Applicants do not wish to be bound by any theoreticalexplanation of the mechanism by which the superior oral bioavailabilityof the present HLCD formulation is achieved.

Antitumor Compositions Comprising HLCD

Yet another preferred embodiment of the claimed invention is anantitumor composition comprising a solution or suspension of a HLCDdissolved or suspended in NMP or NMP containing from about 1.0 mg toabout 40.0 mg HLCD activity per milliliter of solution or suspension andcontaining from about 100 to about 5000 parts of a pharmaceuticallyacceptable carboxylic acid or hydrochloric acid, or phosphoric acid perpart by weight of HLCD. The inventors prefer to use 1000 to 2000 partsby weight of a pharmaceutically acceptable carboxylic acid and/ormineral acid per part by weight of HLCD. For the purpose of thisinvention, examples of carboxylic acid that can be used in thisinvention are tartaric, lactic, ascorbic, gluconic, citric, succinic,fumaric, or maleic acids and examples of mineral acids useful in thisinvention are hydrochloric acid or phosphoric acid.

Another embodiment of this invention is an antitumor compositioncomprising of a solution or suspension of HLCD dissolved or suspended inNMP in the presence of a pharmaceutically acceptable acid, wherein saidsolution or suspension further comprises polyethylene glycol.

Another embodiment of this invention is an antitumor compositioncomprising a solution or suspension of HLCD dissolved in NMP in thepresence of a pharmaceutically acceptable acid, wherein said solution orsuspension further comprises polyethylene glycol and ethyl alcohol orbenzyl alcohol or the solution or suspension further comprises ethylalcohol and benzyl alcohol.

Another embodiment of this invention is an antitumor compositioncomprising a solution or suspension of HLCD dissolved or suspended inNMP in the presence of a pharmaceutically acceptable acid, wherein saidsolution or suspension further comprises polyethylene glycol, andpolysorbate-80.

Another embodiment of this invention is an antitumor compositioncomprising a solution or suspension of HLCD dissolved in NMP in thepresence of a pharmaceutically acceptable acid, wherein said solution orsuspension further comprises polyethylene glycol, ethyl alcohol orbenzyl alcohol (or ethyl alcohol and benzyl alcohol) and polysorbate-80.

Another embodiment of this invention is an antitumor compositioncomprising a solution or suspension of HLCD dissolved or suspended inNMP in the presence of a pharmaceutically acceptable acid, wherein saidsolution or suspension further comprises polyethylene glycol,polysorbate-80 and taurocholic acid or a pharmaceutically acceptablesalt thereof.

Yet another embodiment of this invention is wherein the solution orsuspension of antitumor composition contains for each part by weight ofHLCD, 1,000-10,000 parts by weight of NMP, 100-5,000 parts by weight ofa pharmaceutically acceptable acid, 1-10 parts by weight of taurocholicacid or a pharmaceutically acceptable salt thereof, 1,000-10,000 partsby weight of polyethylene glycol (PEG-300 and/or PEG-400). An additionalembodiment is wherein said acid is an carboxylic acid and the inventorsprefer citric acid.

Another embodiment of the claimed invention is the antitumor compositionfurther comprising a lower alcohol. Many different alcohols would beeffective in this invention, but the inventors prefer to use ethanol ora combination of ethanol and benzyl alcohol. Another embodiment of theclaimed invention is an antitumor composition further comprised ofglycerin as a co-solvent.

Yet another embodiment of this invention is an antitumor compositioncomprising a solution or suspension of HLCD dissolved or suspended inNMP in the presence of a pharmaceutically acceptable acid preferablycitric acid and or hydrochloric or phosphoric acid, polyethylene glycol(PEG-300 and/or PEG-400), polysorbate-80, ethanol, and glycerin.

An additional embodiment of this invention is wherein said solution orsuspension contains for each part by weight of HLCD, 1,000-10,000 partsby weight of NMP, 1,000-5,000 parts by weight of a pharmaceuticallyacceptable acid, 1-10 parts by weight of taurocholic acid or apharmaceutically acceptable salt thereof, 1,000-10,000 parts by weightof polyethylene glycol, 0.1-2.0 parts by weight of glycerin, 1,000-5,000parts by weight of ethanol.

Yet another embodiment of this invention is an antitumor compositioncomprising a solution or suspension of HLCD dissolved or suspended inNMP in the presence of a pharmaceutically acceptable acid, wherein saidsolution or suspension further comprises ethyl alcohol or ethyl alcoholand benzyl alcohol, and polyethylene glycol.

As a more preferred embodiment for this antitumor composition, thepharmaceutically acceptable acid is citric acid, the polyethylene glycolis PEG-400, the lower alcohol is ethanol and the surfactant ispolysorbate-80.

Another embodiment of this invention, is an antitumor compositioncomprising a solution or suspension of about 1.0 mg to about 150.0 mg ofHLCD dissolved or suspended in 1,000-10,000 parts by weight of NMP inthe presence of about 100 to 5000 parts by weight of a pharmaceuticallyacceptable-organic carboxylic acid. This antitumor composition furthercomprises about 1,000-10,000 parts by weight of polyethylene glycol,about 1,000 to 5,000 parts of a pharmaceutically acceptable alcohol.

More preferred for this antitumor composition is when the acid is citricacid, the polyethylene glycol is PEG-400, the alcohol is ethanol and thesurfactant is polysorbate-80.

Another embodiment of this invention is an antitumor compositioncomprising a solution or suspension about 1.0 mg to about 150.0 mg ofHLCD dissolved or suspended in 1,000 to 10,000 parts of NMP in thepresence of 100 to 5,000 parts of a pharmaceutically acceptablecarboxylic acid. This solution or suspension further comprises about1,000 to 5,000 parts of a pharmaceutically acceptable alcohol 1,000 to10,000 part of polyethylene glycol, and 1,000 to 10,000 parts ofpolysorbate-80.

More specifically for this antitumor composition, the acid is citricacid, the alcohol is ethanol, and the polyethylene glycol is PEG-400.

Another embodiment of this invention is an antitumor compositioncomprising a solution or suspension of 1.0 mg to about 150.0 mg of HLCDdissolved or suspended in 1,000 to 10,000 parts of NMP, wherein thissolution or suspension further comprises about 1,000 to 10,000 partspolyoxyethylated castor oil, about 1,000 to 5,000 parts by weight ethylalcohol, and about 1,000 to 5,000 parts citric acid, 1,000 to 10,000parts of polyethylene glycol, and 1,000 to 10,000 parts ofpolysorbate-80.

In a more preferred embodiment, HLCD is solubilized or suspended in amanner suitable for clinical use by forming a solution or suspension of1.0 to 40.0 mg of HLCD per 1 ml in a vehicle comprising 1,000 to 10,000parts by weight of NMP, ethyl alcohol 1,000 to 5,000 parts by weight,benzyl alcohol-1,000 to 5,000 parts by weight, citric acid 1,000 to5,000 parts by weight, polyethylene glycol (PEG-300 or PEG-400) 1,000 to10,000 parts by weight, and polysorbate-80 (Tween-80) 1,000 to 10,000parts. While either polyethylene glycol (PEG-300 or PEG-400) would beeffective in this embodiment, the inventors prefer to employ PEG-400.

This preferred embodiment of a HLCD composition is summarized in Table 6as follows:

                  TABLE 6    ______________________________________    COMPONENT PARTS BY WEIGHT FOR    PARENTERAL OR ORAL FORMULATIONS OF HLCD    Ingredients         Parts by Weight    ______________________________________    HLCD                1.0 to 40.0    .sup.(1) EtOH       1,000 to 5,000    .sup.(1) Benzyl Alcohol                        1,000 to 5,000    Acid                100 to 5,000    PEG 400             1,000 to 10,000    NMP                 1,000 to 10,000    .sup.(1) Cremaphor-EL                        1,000 to 10,000    .sup.(2) Glycerin   0.5 to 2.5    .sup.(2) Taurocholic Acid                        1 to 10    Polysorbate 80      1,000 to 10,000    (Tween-80)    ______________________________________     .sup.(1) optional additions individually or in any combination to oral or     parenteral HLCD formulations     .sup.(2) used in oral formulations only

Another more preferred parenteral formulation comprises HLCD formulatedfor dilution prior to parenteral administration made of 1 to 40 mg ofHLCD in 1 ml of solvents including 1,000 to 10,000 parts by weight ofHLCD of Cremaphor EL (polyoxyethylated castor oil), 1,000 to 5,000 partsethyl alcohol, NMP 1,000 to 10,000 parts, and citric acid 1,000 to 5,000parts.

Yet another embodiment of this invention for oral administration to apatient with cancer is the HLCD dissolved or suspended in NMP in thepresence of a pharmaceutically acceptable acid.

A further embodiment of this invention is the claimed composition andmethod of administering the composition by encapsulating the claimedformulations within a hard gelatin capsule. Yet another embodiment ofthe claimed composition and method of administering the composition isencapsulating the claimed formulations within a soft gelatin capsule orhard gelatin capsule. One of ordinary skill in the art will know thatany of the claimed formulations adapted for oral administration can beused as the fill for the soft or hard gelatin capsule.

A more specific embodiment of the claimed invention is an oralformulation of HLCD in hard or soft gelatin capsules (comprised ofgelatin/glycerin/sorbitol/purifiers) containing 1.0 to 40.0 mg of HLCDper milliliter in a solution or suspension comprising citric acid 1,000to 5,000 parts by weight, glycerin 0.5 to 2.5 parts by weight,polyethylene glycol (molecular weight 300 to 400) 1,000 to 10,000 partsby weight, ethyl alcohol 1,000 to 5,000 parts by weight, PEG-400 1,000to 10,000 parts by weight, polysorbate-80 1,000 to 10,000 parts byweight, and 1,000 to 10,000 parts NMP.

Another preferred oral formulation will include the addition oftaurocholic acid 1 to 10 parts by weight. The soft gelatin capsules mayalso be composed of any of a number of compounds used for this purposeincluding, for example, a mixture of gelatin, glycerin, sorbitol, andparabens.

Table 7 below indicates parts by weight of different components to beincluded in the oral formulation to be administered in capsules. Severalcomponents are marked with an "**" which denotes that the components are"optional." For the purpose of this invention, inclusion of thesecomponents depends on a variety of different factors; i.e. type ofcancer the patient has, pretreated previously, etc.

                  TABLE 7    ______________________________________    COMPONENT PARTS BY WEIGHT FOR ORAL    FORMULATION OF HLCD    Ingredients          Parts by Weight    ______________________________________    HLCD                 1 to 40    NMP                  1,000 to 10,000    Citric Acid          1,000 to 5,000    EtOH                 1,000 to 5,000    Polysorbate-80 (Tween-80)                         1,000 to 10,000    PEG-400              1,000 to 10,000    Glycerin**           0.5 to 2.5    Taurocholic Acid**   1 to 10    ______________________________________

Clinicians will administer HLCD in these formulations to human patientswith cancer according to schedules which maximize its potentialantitumor effects and diminish its potential toxic side effects. Exceptat extremely high doses which produce high plasma concentrations of thedrug, the antitumor activity of HLCD can be increased by increasing theduration of exposure (time dependent) rather than increasing the dose(dose dependent) of the drug. Increased antitumor effects associatedwith increasing the duration of exposure is most likely related to thepredominant S-phase mode of antitumor activity of HLCD. HLCD areS-phase-active agents therefore, the greatest antitumor effect in humanswill likely be observed with prolonged infusion or closely spacedrepetitive administration schedules. Such schedules of administrationwould expose more cycling tumor cells to the drug and increase thefrequency of exposure of the tumor-cells in S-phase to sufficientlytoxic levels of the drug.

A further embodiment of this invention is that the claimed HLCDantitumor composition can be used to treat a variety of different cancertypes. The claimed formulations and compositions of this invention maybe used in treatment of a number of tumors (cancers) including, withoutlimitation, human cancers of the lung, breast, colon, prostate,melanoma, pancreas, stomach, liver, brain, kidney, uterus, cervix,ovaries, and urinary tract.

In many cases, the site and type of tumor to be treated will influencethe preferred route of administration and therapeutic regimen to beapplied. Consequently, although the formulations of the invention may bemost usually administered by intravenous injection, infusion or orally,these formulations may also can be delivered directly into the tumorsite or by other methods designed to target the drug directly to thetumor site. For example, in patients with malignant pleural effusion,the intrapleural route may be preferred; in patients with poor venousaccess the subcutaneous route of administration may be preferred; inpatients with primary or metastatic cancer involving the brain ornervous system, the intracisternal or intrathecal route ofadministration may be most advantageous; in patients with malignantascites secondary to cancer, one may select intraperitonealadministration; and in patients with bladder cancer directintravesicular instillation may be most advantageous. Similarly, intumors of the skin, the formulation may be topically applied. An oralformulation is also taught for use where suitable for patients takingthe medication outside of the hospital or clinic.

An additional embodiment of this invention is a HLCD solution orsuspension comprising HLCD dissolved or suspended in NMP, in thepresence of a pharmaceutically acceptable acid and this solution orsuspension is prepared for oral, intrapleural, intrathecal,subcutaneous, intracisternal, intravesicular, intraperitoneal, topicalor intravenous administration to a patient with cancer.

A further embodiment of claimed HLCD is a method of treatment of cancerin humans with convergent therapy or combination therapy. This methoduses HLCD dissolved in NMP, in the presence of pharmaceuticallyacceptable acid and co-administers it with additional drugs selectedfrom the group consisting of, but not limited to, carmustine,azathioprine, cis-platinum, carboplatin, iproplatin, cyclophosphamide,ifosfamide, etoposide, ara-C, doxorubicin, daunorubicin, nitrogenmustard, 5-fluorouracil, bleomycin, mitomycin-C, fluoxymesterone,mechlorethamine, teniposide, hexamethylmelamine, leucovorin, melphelan,methotrexate, mercaptopurine, mitoxantrone, BCNU, CCNU, procarbazine,vincristine, vinblastine, vindesine, thioTEPA, amsacrine, G-CSF, GM-CSF,erythropoietin, γ-methylene-10-deazaaminopterin orγ-methylene-10-ethyl-10-deazaaminopterin, taxol, and 5-azacytidine. Forthe purpose of this invention, the terms convergent, co-administered,and combination are used interchangeably.

HLCD dissolved or suspended in NMP with or without the describedcombination of other formulation excipients that have been taught in theforegoing section have additional utility when administered parenterallyusing a prolonged schedule of administration. To increase the utility ofHLCD for parenteral infusions, the HLCD parenteral compositions may bediluted with an appropriate volume of an aqueous vehicle to aconcentration of about 0.001 mg/ml to 1.0 mg/ml of HLCD activity.

A further embodiment of the claimed invention is a solution of any ofthe claimed HLCD compositions and formulations for administration to apatient with cancer upon dilution with a sterile aqueous parenteralvehicle. For the purposes of this invention, parenteral aqueous vehiclessuitable for dilution include dextrose 5 to 10% in water, 0.9% NaCl inwater with or without 5% or 10% Dextrose, 0.45% NaCl in water with orwithout 5% or 10% Dextrose, Lactated Ringer's Solution, 3% NaCl in waterwith or without 5% to 10% Dextrose, water USP for injection or sterilelipid formulations, such as intralipid, used for parenteral nutritionalsupport for cancer patients.

This invention is also directed to a solution or suspension comprisingan A-ring substituted camptothecin having a water solubility of 5micrograms or less than 5 micrograms per milliliter wherein thesubstituted camptothecin is dissolved or suspended in an effectiveamount of N-methyl-2-pyrrolidinone. A-ring substitutions include anysubstitutions on the A-ring but the inventors prefer to employsubstitutions at position 9, 10 or 11 of the A-ring.

This invention is also directed to a solution or suspension comprisingan B-ring substituted camptothecin having a water solubility of 5micrograms or less than 5 micrograms per milliliter wherein thesubstituted camptothecin is dissolved or suspended in an effectiveamount of N-methyl-2-pyrrolidinone. B-ring substitutions include anysubstitutions on the B-ring but the inventors prefer to employsubstitutions at position 7 of the B-ring.

This invention is also directed to a solution or suspension comprising asubstituted camptothecin having a water solubility of 5 micrograms orless than 5 micrograms per milliliter wherein the camptothecin hassubstitutions on the A-ring and on the B-ring and wherein thesubstituted camptothecin is dissolved or suspended in an effectiveamount of N-methyl-2-pyrrolidinone. This invention includessubstitutions at all possible locations on the A- and B-ring but theinventors prefer substitutions at position 7 of the B-ring and atpositions 9, 10, and/or 11 of the A-ring.

This invention is also directed to a solution or suspension comprisingan B-ring substituted camptothecin having a water solubility of 5micrograms or less than 5 micrograms per milliliter wherein thesubstituted camptothecin is dissolved or suspended in an effectiveamount of N-methyl-2-pyrrolidinone.

All of the embodiments outlined below apply to A-ring substitutedcamptothecins, B-ring substituted camptothecins and to substitutedcamptothecins containing both A-ring and B-ring substitutions. For thisinvention, "dissolved" and "suspended" have regular meanings known toone of ordinary skill in this art.

The above solution or suspension may further contain a pharmaceuticallyacceptable acid wherein this acid can be a carboxylic acid selected fromthe group consisting of acetic acid, citric acid, fumaric acid, maleicacid, ascorbic acid, gluconic acid, and lactic acid. The above solutionor suspension may further contain a pharmaceutically acceptable acidwherein this acid can be a mineral acid selected from a group consistingof hydrochloric acid and phosphoric acid. Additionally, thepharmaceutically acceptable acid in the above solution or suspension maybe selected from the group consisting of acetic acid, citric acid,fumaric acid, maleic acid, ascorbic acid, phosphoric acid, gluconicacid, lactic acid, and hydrochloric acid. Another embodiment of thisinvention is the above solution or suspension prepared and sterilizedfor oral, intrapleural, intrathecal, intracisternal, intravesicular,intraperitoneal, topical or intravenous administration to a patient withcancer. Yet another embodiment of this invention is this solution orsuspension is encapsulated within a hard gelatin capsule or a softgelatin capsule.

Another embodiment of this invention is the above solution or suspensionmay further contain a lower alcohol selected from the group consistingof ethanol and benzyl alcohol. The above solution or suspension may alsocontain a polyethylene glycol selected from a group consisting ofPEG-300 and PEG-400. And the above solution or suspension may furthercontain a non-ionic surfactant. The inventors prefer to employpolysorbate-80 (Tween-80) but most surfactants are suitable. Anotherembodiment of this invention is the above solution or suspensionprepared and sterilized for oral, intrapleural, intrathecal,intracisternal, intravesicular, intraperitoneal, topical or intravenousadministration to a patient with cancer. Yet another embodiment of thisinvention is this solution or suspension is encapsulated within a hardgelatin capsule or a soft gelatin capsule.

Another embodiment of this invention is the above solution or suspensioncomprising an A-ring substituted or B-ring substituted or A- and B-ringsubstituted camptothecin and N-methyl-2-pyrrolidinone may also furthercontain a polyethylene glycol and a non-ionic surfactant. Anotherembodiment of this invention is the above solution or suspensionprepared and sterilized for oral, intrapleural, intrathecal,intracisternal, intravesicular, intraperitoneal, topical or intravenousadministration to a patient with cancer. Yet another embodiment of thisinvention is this solution or suspension is encapsulated within a hardgelatin capsule or a soft gelatin capsule.

The above solution or suspension comprising an A-ring substituted orB-ring substituted or A- and B-ring substituted camptothecin andN-methyl-2-pyrrolidinone may also contain a lower alcohol, polyethyleneglycol and a non-ionic surfactant. Another embodiment of this inventionis the above solution or suspension prepared and sterilized for oral,intrapleural, intrathecal, intracisternal, intravesicular,intraperitoneal, topical or intravenous administration to a patient withcancer. Yet another embodiment of this invention is this solution orsuspension is encapsulated within a hard gelatin capsule or a softgelatin capsule.

Another embodiment of this invention is the above solution or suspensioncomprising an A-ring substituted or B-ring substituted or A- and B-ringsubstituted camptothecin and N-methyl-2-pyrrolidinone may also furthercontain 1 to 10 parts by weight of taurocholic acid or apharmaceutically acceptable salt thereof. Another embodiment of thisinvention is the above solution or suspension prepared and sterilizedfor oral, intrapleural, intrathecal, intracisternal, intravesicular,intraperitoneal, topical or intravenous administration to a patient withcancer. Yet another embodiment of this invention is this solution orsuspension is encapsulated within a hard gelatin capsule or a softgelatin capsule.

Another embodiment of this invention is the above solution or suspensioncomprising an A-ring substituted or B-ring substituted or A- and B-ringsubstituted camptothecin, N-methyl-2-pyrrolidinone, and acid may alsofurther contain 1 to 10 parts by weight of taurocholic acid or apharmaceutically acceptable salt thereof. Another embodiment of thisinvention is the above solution or suspension prepared and sterilizedfor oral, intrapleural, intrathecal, intracisternal, intravesicular,intraperitoneal, topical or intravenous administration to a patient withcancer. Yet another embodiment of this invention is this solution orsuspension is encapsulated within a hard gelatin capsule or a softgelatin capsule.

Yet another embodiment of this invention is the above solution orsuspension comprising an A-ring substituted or B-ring substituted or A-and B-ring substituted camptothecin, N-methyl-2-pyrrolidinone, acid, andlower alcohol may also further contain 1 to 10 parts by weight oftaurocholic acid or a pharmaceutically acceptable salt thereof. Anotherembodiment of this invention is the above solution or suspensionprepared and sterilized for oral, intrapleural, intrathecal,intracisternal, intravesicular, intraperitoneal, topical or intravenousadministration to a patient with cancer. Yet another embodiment of thisinvention is this solution or suspension is encapsulated within a hardgelatin capsule or a soft gelatin capsule.

Yet another embodiment of this invention is the above solution orsuspension comprising an A-ring substituted or B-ring substituted or A-and B-ring substituted camptothecin, N-methyl-2-pyrrolidinone, acid, andpolyethylene glycol may also further contain 1 to 10 parts by weight oftaurocholic acid or a pharmaceutically acceptable salt thereof. Anotherembodiment of this invention is the above solution or suspensionprepared and sterilized for oral, intrapleural, intrathecal,intracisternal, intravesicular, intraperitoneal, topical or intravenousadministration to a patient with cancer. Yet another embodiment of thisinvention is this solution or suspension is encapsulated within a hardgelatin capsule or a soft gelatin capsule.

Another embodiment of this invention is the above solution or suspensioncomprising an A-ring substituted or B-ring substituted or A- and B-ringsubstituted camptothecin, N-methyl-2-pyrrolidinone, acid, and non-ionicsurfactant may also further contain 1 to 10 parts by weight oftaurocholic acid or a pharmaceutically acceptable salt thereof. Anotherembodiment of this invention is the above solution or suspensionprepared and sterilized for oral, intrapleural, intrathecal,intracisternal, intravesicular, intraperitoneal, topical or intravenousadministration to a patient with cancer. Yet another embodiment of thisinvention is this solution or suspension is encapsulated within a hardgelatin capsule or a soft gelatin capsule.

Another embodiment of this invention is the above solution or suspensioncomprising an A-ring substituted or B-ring substituted or A- and B-ringsubstituted camptothecin, N-methyl-2-pyrrolidinone, acid, polyethyleneglycol and non-ionic surfactant may also further contain 1 to 10 partsby weight of taurocholic acid or a pharmaceutically acceptable saltthereof. Another embodiment of this invention is the above solution orsuspension prepared and sterilized for oral, intrapleural, intrathecal,intracisternal, intravesicular, intraperitoneal, topical or intravenousadministration to a patient with cancer. Yet another embodiment of thisinvention is this solution or suspension is encapsulated within a hardgelatin capsule or a soft gelatin capsule.

Another embodiment of this invention is the above solution or suspensioncomprising an A-ring substituted or B-ring substituted or A- and B-ringsubstituted camptothecin, N-methyl-2-pyrrolidinone, acid, polyethyleneglycol, non-ionic surfactant and lower alcohol may also further contain1 to 10 parts by weight of taurocholic acid or a pharmaceuticallyacceptable salt thereof. Another embodiment of this invention is theabove solution or suspension prepared and sterilized for oral,intrapleural, intrathecal, intracisternal, intravesicular,intraperitoneal, topical or intravenous administration to a patient withcancer. Yet another embodiment of this invention is this solution orsuspension is encapsulated within a hard gelatin capsule or a softgelatin capsule.

Another embodiment of this invention is the above solution or suspensioncomprising an A-ring substituted or B-ring substituted or A- and B-ringsubstituted camptothecin and N-methyl-2-pyrrolidinone wherein thissolution or suspension contains from about 1.0 mg to about 40.0 mgactivity per ml of solution or suspension of the substitutedcamptothecin having a water solubility of less than 5 micrograms per mlof water. Another embodiment of this invention is the above solution orsuspension prepared and sterilized for oral, intrapleural, intrathecal,intracisternal, intravesicular, intraperitoneal, topical or intravenousadministration to a patient with cancer. Yet another embodiment of thisinvention is this solution or suspension is encapsulated within a hardgelatin capsule or a soft gelatin capsule.

Yet another embodiment of this invention is an antitumor compositioncomprising a solution or suspension containing an A-ring substitutedcamptothecin or containing a B-ring substituted camptothecin orcontaining both an A-ring substituted and a B-ring substitutedcamptothecin having a water solubility of 5 micrograms or less than 5micrograms per milliliter dissolved or suspended inN-methyl-2-pyrrolidinone wherein said composition contains from about 1mg to about 40 mg camptothecin activity per ml and contains about 1,000to 10,000 parts by weight of N-methyl-2-pyrrolidinone.

For this invention "substituted camptothecin" applies to substitutionsin the A-ring or substitutions in the B-ring or substitutions in boththe A-ring and the B-ring. This invention includes substitutions at allpossible locations on the A- and B-ring but the inventors prefersubstitutions at position 7 of the B-ring and at positions 9, 10, and/or11 of the A-ring.

The antitumor composition comprising a substituted camptothecin andN-methyl-2-pyrrolidinone may also contain from about 100 to about 5,000parts by weight of a pharmaceutically acceptable acid per part by weightof the substituted camptothecin.

The antitumor composition comprising a substituted camptothecin,N-methyl-2-pyrrolidinone, and acid may further contain 1,000 to 10,000parts by weight of polyethylene glycol selected from the groupconsisting of PEG-300 and PEG-400.

The antitumor composition comprising a substituted camptothecin,N-methyl-2-pyrrolidinone, and acid may further contain a non-ionicsurfactant. Many different non-ionic surfactants are available but theinventors prefer to employ polysorbate-80 (Tween-80).

The antitumor composition comprising a substituted camptothecin,N-methyl-2-pyrrolidinone, and acid may further contain a lower alcoholselected from the group consisting of ethanol and benzyl alcohol.

The antitumor composition comprising a substituted camptothecin,N-methyl-2-pyrrolidinone, and acid may further a lower alcohol,polyethylene glycol, and a non-ionic surfactant.

The antitumor composition comprising a substituted camptothecin,N-methyl-2-pyrrolidinone, and acid and further containing a loweralcohol, polyethylene glycol, and a non-ionic surfactant wherein saidacid is citric acid, wherein said lower alcohol is selected from thegroup consisting of ethanol and benzyl alcohol, wherein saidpolyethylene glycol is selected from the group consisting of PEG-300 andPEG-400, and wherein said surfactant is polysorbate-80.

Another embodiment of this invention is an antitumor compositioncomprising a solution or suspension containing an A-ring substitutedcamptothecin or containing a B-ring substituted camptothecin orcontaining both an A-ring substituted and a B-ring substitutedcamptothecin having a water solubility of 5 micrograms or less than 5micrograms per milliliter wherein for each part by weight of substitutedcamptothecin said solution or suspension contains 1,000 to 10,000 partsN-methyl-2-pyrrolidinone, 100 to 5,000 parts of a pharmaceuticallyacceptable acid, 1,000 to 10,000 parts by weight of polyethylene glycol,and 1,000 to 5,000 parts of lower alcohol selected from the groupconsisting of ethanol and benzyl alcohol.

Yet another embodiment of this invention is an antitumor compositioncomprising a solution or suspension containing an A-ring substitutedcamptothecin or containing a B-ring substituted camptothecin orcontaining both an A-ring substituted and a B-ring substitutedcamptothecin having a water solubility of 5 micrograms or less than 5micrograms per milliliter wherein for each part by weight of substitutedcamptothecin said solution or suspension contains 1,000 to 10,000 partsN-methyl-2-pyrrolidinone, 100 to 5,000 parts of a pharmaceuticallyacceptable acid, 1,000 to 5,000 parts of a lower alcohol, and 1,000 to10,000 parts of a non-ionic surfactant. This invention also embodies theabove antitumor composition wherein the acid is citric acid, wherein thealcohol is ethanol, and wherein the non-ionic surfactant ispolysorbate-80.

For this invention "substituted camptothecin" applies to substitutionsin the A-ring or substitutions in the B-ring or substitutions in boththe A-ring and the B-ring. This invention includes substitutions at allpossible locations on the A- and B-ring but the inventors prefersubstitutions at position 7 of the B-ring and at positions 9, 10, and/or11 of the A-ring.

This invention further embodies an injectable, sterile solutioncomprising N-methyl-2-pyrrolidinone, and a highly lipophiliccamptothecin derivative having a water solubility of 5 micrograms permilliliter or less, with the general structural formula: ##STR3##wherein R₁, R₂, R₃, R₄ =H, lower alkyl, alkoxy, acyloxy, hydroxy, acyl,halo, amido, or cyano group;

wherein R₁ and R₂ together may represent --X₁ --X₂ --X₃ -- and whereinX₁, X₂, X₃ may be CR₅ R₆, O, S or NR₇ and wherein R₅, R₆, R₇ =H, loweralkyl, alkoxy, acyloxy, hydroxy, acyl, halo, amido, or cyano group;

wherein R₂ and R₃ together may represent --X₁ --X₂ --X₃ -- and whereinX₁, X₂, X₃ may be CR₅ R₆, O, S or NR₇ and wherein R₅, R₆, R₇ =H, loweralkyl, alkoxy, acyloxy, hydroxy, acyl, halo, amido, or cyano group;

wherein R₃ and R₄ together may represent --X₁ --X₂ --X₃ -- and whereinX₁, X₂, X₃ may be CR₅ R₆, O, S or NR₇ and wherein R₅, R₆, R₇ =H, loweralkyl, alkoxy, acyloxy, hydroxy, acyl, halo, amido, or cyano group;

Another embodiment of this invention is the above injectable, sterilesolution comprising N-methyl-2-pyrrolidinone, and a highly lipophiliccamptothecin derivative further containing a pharmaceutically acceptableacid. This acid may be a carboxylic acid selected from the groupconsisting of acetic acid, citric acid, fumaric acid, maleic acid,ascorbic acid, gluconic acid, and lactic acid. Or, this acid may be amineral acid selected from a group consisting of hydrochloric acid andphosphoric acid. The acid may also an acid selected from the groupconsisting of acetic acid, citric acid, fumaric acid, maleic acid,ascorbic acid, phosphoric acid, gluconic acid, lactic acid, andhydrochloric acid.

Yet another embodiment of this invention is the above injectable,sterile solution comprising N-methyl-2-pyrrolidinone, a highlylipophilic camptothecin derivative, and a pharmaceutically acceptableacid further containing a lower alcohol selected from the groupconsisting of ethanol and benzyl alcohol.

Yet another embodiment of this invention is the above injectable,sterile solution comprising N-methyl-2-pyrrolidinone, a highlylipophilic camptothecin derivative, and a pharmaceutically acceptableacid further containing a polyethylene glycol selected from a groupconsisting of PEG-300 and PEG-400.

Yet another embodiment of this invention is the above injectable,sterile solution comprising N-methyl-2-pyrrolidinone, a highlylipophilic camptothecin derivative, and a pharmaceutically acceptableacid further containing a non-ionic surfactant wherein the non-ionicsurfactant is polysorbate-80 (Tween-80).

Yet another embodiment of this invention is the above injectable,sterile solution comprising N-methyl-2-pyrrolidinone, a highlylipophilic camptothecin derivative, and a pharmaceutically acceptableacid further containing a polyethylene glycol and a non-ionicsurfactant.

Yet another embodiment of this invention is the above injectable,sterile solution comprising N-methyl-2-pyrrolidinone, a highlylipophilic camptothecin derivative, and a pharmaceutically acceptableacid further containing a lower alcohol, polyethylene glycol and anon-ionic surfactant.

Another embodiment of this invention is the above injectable sterilesolution comprising an A-ring substituted or B-ring substituted or A-and B-ring substituted camptothecin and N-methyl-2-pyrrolidinone mayalso further contain 1 to 10 parts by weight of taurocholic acid or apharmaceutically acceptable salt thereof. Another embodiment of thisinvention is the above solution or suspension prepared and sterilizedfor oral, intrapleural, intrathecal, intracisternal, intravesicular,intraperitoneal, topical or intravenous administration to a patient withcancer. Yet another embodiment of this invention is this solution orsuspension is encapsulated within a hard gelatin capsule or a softgelatin capsule.

Another embodiment of this invention is the above injectable sterilesolution comprising an A-ring substituted or B-ring substituted or A-and B-ring substituted camptothecin, N-methyl-2-pyrrolidinone, and acidmay also further contain 1 to 10 parts by weight of taurocholic acid ora pharmaceutically acceptable salt thereof. Another embodiment of thisinvention is the above solution or suspension prepared and sterilizedfor oral, intrapleural, intrathecal, intracisternal, intravesicular,intraperitoneal, topical or intravenous administration to a patient withcancer. Yet another embodiment of this invention is this solution orsuspension is encapsulated within a hard gelatin capsule or a softgelatin capsule.

Yet another embodiment of this invention is the above injectable sterilesolution comprising an A-ring substituted or B-ring substituted or A-and B-ring substituted camptothecin, N-methyl-2-pyrrolidinone, acid, andlower alcohol may also further contain 1 to 10 parts by weight oftaurocholic acid or a pharmaceutically acceptable salt thereof. Anotherembodiment of this invention is the above solution or suspensionprepared and sterilized for oral, intrapleural, intrathecal,intracisternal, intravesicular, intraperitoneal, topical or intravenousadministration to a patient with cancer. Yet another embodiment of thisinvention is this solution or suspension is encapsulated within a hardgelatin capsule or a soft gelatin capsule.

Yet another embodiment of this invention is the above injectable sterilesolution comprising an A-ring substituted or B-ring substituted or A-and B-ring substituted camptothecin, N-methyl-2-pyrrolidinone, acid, andpolyethylene glycol may also further contain 1 to 10 parts by weight oftaurocholic acid or a pharmaceutically acceptable salt thereof. Anotherembodiment of this invention is the above solution or suspensionprepared and sterilized for oral, intrapleural, intrathecal,intracisternal, intravesicular, intraperitoneal, topical or intravenousadministration to a patient with cancer. Yet another embodiment of thisinvention is this solution or suspension is encapsulated within a hardgelatin capsule or a soft gelatin capsule.

Another embodiment of this invention is the above injectable sterilesolution comprising an A-ring substituted or B-ring substituted or A-and B-ring substituted camptothecin, N-methyl-2-pyrrolidinone, acid, andnon-ionic surfactant may also further contain 1 to 10 parts by weight oftaurocholic acid or a pharmaceutically acceptable salt thereof. Anotherembodiment of this invention is the above solution or suspensionprepared and sterilized for oral, intrapleural, intrathecal,intracisternal, intravesicular, intraperitoneal, topical or intravenousadministration to a patient with cancer. Yet another embodiment of thisinvention is this solution or suspension is encapsulated within a hardgelatin capsule or a soft gelatin capsule.

Another embodiment of this invention is the above injectable sterilesolution comprising an A-ring substituted or B-ring substituted or A-and B-ring substituted camptothecin, N-methyl-2-pyrrolidinone, acid,polyethylene glycol and non-ionic surfactant may also further contain 1to 10 parts by weight of taurocholic acid or a pharmaceuticallyacceptable salt thereof. Another embodiment of this invention is theabove solution or suspension prepared and sterilized for oral,intrapleural, intrathecal, intracisternal, intravesicular,intraperitoneal, topical or intravenous administration to a patient withcancer. Yet another embodiment of this invention is this solution orsuspension is encapsulated within a hard gelatin capsule or a softgelatin capsule.

Another embodiment of this invention is the above injectable sterilesolution comprising an A-ring substituted or B-ring substituted or A-and B-ring substituted camptothecin, N-methyl-2-pyrrolidinone, acid,polyethylene glycol, non-ionic surfactant and lower alcohol may alsofurther contain 1 to 10 parts by weight of taurocholic acid or apharmaceutically acceptable salt thereof. Another embodiment of thisinvention is the above solution or suspension prepared and sterilizedfor oral, intrapleural, intrathecal, intracisternal, intravesicular,intraperitoneal, topical or intravenous administration to a patient withcancer. Yet another embodiment of this invention is this solution orsuspension is encapsulated within a hard gelatin capsule or a softgelatin capsule.

Another embodiment of this invention is the above injectable sterilesolution comprising an A-ring substituted or B-ring substituted or A-and B-ring substituted camptothecin and N-methyl-2-pyrrolidinone whereinthis solution or suspension contains from about 1.0 mg to about 40.0 mgactivity per ml of solution or suspension of the substitutedcamptothecin having a water solubility of less than 5 micrograms per mlof water. Another embodiment of this invention is the above solution orsuspension prepared and sterilized for oral, intrapleural, intrathecal,intracisternal, intravesicular, intraperitoneal, topical or intravenousadministration to a patient with cancer. Yet another embodiment of thisinvention is this solution or suspension is encapsulated within a hardgelatin capsule or a soft gelatin capsule.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

In its preferred embodiments, this invention involves preparation anduse of novel HLCD solutions or suspensions as described below.

EXAMPLES

The following examples illustrate selected modes for carrying out theclaimed invention and are not to be construed as limiting thespecification and claims in any way. These examples are provided so asto enable those in ordinary skill in the art to make the compositions ofthis invention. These examples are not intended to limit the scope ofwhat the inventors regard as the invention. Efforts have been made toensure accuracy with respect to numbers used to characterize themeasured conditions; however, some experimental errors and deviationsmay be present.

Maintaining an acidic pH (3 to 5) in the HLCD solution or suspension isparticularly important to reduce the slow conversion of HLCD lactone tothe E-ring-hydrolyzed carboxylate, which occurs at physiological pH. Atequilibrium under physiologic pH, the ratio of the open-ring(carboxylate) form to lactone species increases. Hydrolysis of the HLCDlactone ring will be substantially reduced if the drug is kept in anacidic environment. Some of the unpredictable toxicity seen in earlierclinical trials using SCPT may have been due to the in vivo formation ofgreater amounts of the lactone form of camptothecin, which is 10-foldmore toxic than SCPT in mice. The lactone form of HLCD, as incamptothecin, is less water soluble than the carboxylate E-ring form.When early clinical trials were first conducted with camptothecin usingsodium hydroxide, the significance of maintaining the closed lactonering for uniform efficacy and safety in treating patients with cancerwas poorly understood. The early reported unpredictable clinicaltoxicities associated with camptothecin administration may have beenexacerbated by the sodium hydroxide formulation which promotes theformation of the carboxylate form, and by the relative lack ofunderstanding of the significance of the lactone form of camptothecin asit relates to antitumor activity.

Example 1 Highly Lipophilic Camptothecin Derivative-NMP Formulation forInjection or Infusion

For injection or infusion into aqueous body fluids, a formulationcontains from about 1.0 to about 40.0 parts by weight of HLCD in 1,000to 10,000 parts by weight of NMP in a vehicle comprising between about1,000 to about 5,000 part by weight of an acceptable alcohol, about1,000 to about 10,000 parts by weight of polyethylene glycol, and about1,000 to about 10,000 parts of a non-ionic surfactant. Suitable alcoholsinclude dehydrated ethyl alcohol or benzyl alcohol, or combination ofethyl alcohol and benzyl alcohol. Suitable polyethylene glycols, includepolyethylene glycol 300, polyethylene glycol 400, and polypropyleneglycol. Suitable non-ionic surfactants include polysorbate-80(Tween-80). In a preferred embodiment, the formulation of HLCD issupplied as an intravenous injectable in a vial comprising a solution ofdrug in a vehicle comprising ethyl alcohol and/or benzyl alcohol, citricacid, polyethylene glycol 400, and polysorbate (Tween 80).

Example 2 Highly Lipophilic Camptothecin Derivative-NMP Formulation #2

A second formulation contains from about 1.0 to about 40.0 parts byweight of HLCD in 1,000 to 10,000 part of NMP further comprising betweenabout 1,000 to 5,000 parts of an alcohol, about 100 to 5,000 parts byweight of an pharmaceutically acceptable acid and about 1,000 to 10,000parts of polyethylene glycol 400. Suitable alcohols include ethylalcohol, and benzyl alcohol. Suitable acids for this formulation includecitric acid, hydrochloric acid or phosphoric acid. In a preferredembodiment 1 to about 20 parts of HLCD by weight is formulated in 1,000to 10,000 parts of NMP, 1,000 to 10,000 parts of PEG-400, 1,000 to 5,000parts by weight ethyl alcohol, and 1,000 to 5,000 parts of citric acidby weight.

Example 3 Highly Lipophilic Camptothecin Derivative-NMP Oral Formulation

An oral formulation of HLCD in soft gelatin capsules (comprised ofgelatin/glycerin/sorbitol/purifiers) containing from about 1.0 part toabout 40.0 parts by weight of HLCD dissolved in 1,000 to 10,000 parts ofNMP, citric acid 1,000 to about 5,000 parts by weight, glycerin 0.5 to2.5 parts by weight, and polyethylene glycol (PEG-300 or PEG-400) 1,000to 10,000 parts by weight, ethyl alcohol 1,000 to 5,000 parts by weight,and taurocholic acid 1 to 10 parts by weight. The soft gelatin capsulesmay also be composed of any of a number of compounds used for thispurpose including for example, a mixture of gelatin, glycerin, sorbitol,and parabens.

Example 4 7-Ethyl Camptothecin-NMP Formulation

A formulation containing from about 1.0 to about 40.0 parts by weight of7-ethyl camptothecin in 1,000 to 10,000 part of NMP further comprisingbetween about 1,000 to 5,000 parts of an alcohol, about 100 to 5,000parts by weight of an pharmaceutically acceptable acid and about 1,000to 10,000 parts of polyethylene glycol 400. Suitable alcohols includeethyl alcohol, and benzyl alcohol. Suitable acids for this formulationinclude citric acid, hydrochloric acid or phosphoric acid. In apreferred embodiment 1 to about 20 parts of 7-ethyl camptothecin byweight is formulated in 1,000 to 10,000 parts of NMP, 1,000 to 10,000parts of PEG-400, 1,000 to 5,000 parts by weight ethyl alcohol, and1,000 to 5,000 parts of citric acid by weight.

Example 5 7-Ethyl-10-Hydroxy Camptothecin-NMP Formulation

A formulation containing from about 1.0 to about 40.0 parts by weight of7-ethyl-lo-hydroxy camptothecin in 1,000 to 10,000 part of NMP furthercomprising between about 1,000 to 5,000 parts of an alcohol, about 100to 5,000 parts by weight of an pharmaceutically acceptable acid andabout 1,000 to 10,000 parts of polyethylene glycol 400. Suitablealcohols include ethyl alcohol, and benzyl alcohol. Suitable acids forthis formulation include citric acid, hydrochloric acid or phosphoricacid. In a preferred embodiment 1 to about 20 parts of7-ethyl-10-hydroxy camptothecin by weight is formulated in 1,000 to10,000 parts of NMP, 1,000 to 10,000 parts of PEG-400, 1,000 to 5,000parts by weight ethyl alcohol, and 1,000 to 5,000 parts of citric acidby weight.

Example 6 10,11-Methylenedioxy Camptothecin-NMP Formulation

A formulation containing from about 1.0 to about 40.0 parts by weight of10,11-methylenedioxy camptothecin in 1,000 to 10,000 part of NMP furthercomprising between about 1,000 to 5,000 parts of an alcohol, about 100to 5,000 parts by weight of an pharmaceutically acceptable acid andabout 1,000 to 10,000 parts of polyethylene glycol 400. Suitablealcohols include ethyl alcohol, and benzyl alcohol. Suitable acids forthis formulation include citric acid, hydrochloric acid or phosphoricacid. In a preferred embodiment 1 to about 20 parts of10,11-methylenedioxy by weight is formulated in 1,000 to 10,000 parts ofNMP, 1,000 to 10,000 parts of PEG-400, 1,000 to 5,000 parts by weightethyl alcohol, and 1,000 to 5,000 parts of citric acid by weight.

Example 7 10-Bromo Camptothecin-NMP Formulation

A formulation containing from about 1.0 to about 40.0 parts by weight of10-bromo camptothecin in 1,000 to 10,000 part of NMP further comprisingbetween about 1,000 to 5,000 parts of an alcohol, about 100 to 5,000parts by weight of an pharmaceutically acceptable acid and about 1,000to 10,000 parts of polyethylene glycol 400. Suitable alcohols includeethyl alcohol, and benzyl alcohol. Suitable acids for this formulationinclude citric acid, hydrochloric acid or phosphoric acid. In apreferred embodiment 1 to about 20 parts of 10-bromo camptothecin byweight is formulated in 1,000 to 10,000 parts of NMP, 1,000 to 10,000parts of PEG-400, 1,000 to 5,000 parts by weight ethyl alcohol, and1,000 to 5,000 parts of citric acid by weight.

Example 8

Use of N-methyl-2-pyrrolidinone (NMP) in the Formulations

One of the key discoveries in the present invention is the unexpectedlyhigh solubility of HLCD in N-methyl-2-pyrrolidinone (NMP).N-Methyl-2-pyrrolidinone is an organic liquid excipient and is alsoknown as 1-methylpyrrolidinone, N-methyl-2-pyrrolidinone,1-methyl-5-pyrrolidinone, methylpyrrolidinone, N-methyl pyrrolidinone,methylpyrrolidinone, N-methylpyrrolidone, N-methyl-2-pyrrolidone,M-pyrol, and NMP.

NMP exhibits a high degree of physiologic safety in mammals with thefollowing LD50 values: (rat) oral--7000 mg/kg, intraperitoneal--2472mg/kg, intravenous--2266 mg/kg, (mice) oral--7725 mg/kg,intraperitoneal--4429 mg/kg, intravenous--3605 mg/kg, (rabbit)skin--8000 mg/kg (Registry of Toxic Effects of Chemical Substances,1983-84 Supplement, Page 1628). NMP has been used to formulate etoposide(Etoposide, U.S. Pat. No. 4,772,589) and acridine derivatives (M-AMSA,U.S. Pat. No. 5,034,397). Etoposide and acridine derivatives (1) areanticancer drugs; (2) are chemically unrelated to HLCD; (3) are morewater soluble than HLCD; and (4) exert their antitumor effects by vastlydifferent mechanisms than HLCD.

NMP has also been used for oral formulations of the antibioticclarithromycin (Clarithromycin, WO patent #9,014,094) and other drugs.NMP is a key excipient of the instant invention which allows anexceptionally high degree of drug solubility of HLCD (range 1 mg/ml to40 mg/ml)in NMP as a solution or suspension. An HLCD solution comprisingNMP with or without other combinations of excipients described herein,which can be diluted with a parenteral vehicle such as sterileinjectable water USP, 5% Dextrose solution for injection USP or 0.9%sodium chloride solution for injection USP, such that the amount of HLCDdissolved in the diluted infusion is from about 0.001 mg/ml to about 1.0mg/ml, is taught in the present invention. The inventors have discoveredthat HLCD show remarkably high solubility in NMP compared to othercommon pharmaceutical solvents such as water, ethanol, benzyl alcohol,propylene glycol, PEG 300, PEG 400, dimethylisosorbide ordimethylacetamide (Table 5). This high solubility of HLCD in NMP makesNMP a unique and highly useful pharmaceutical solvent for making usefulsolutions or suspensions of HLCD.

    ______________________________________    Solubility of Camptothecin in Various Solvents    Solvent             Concentration, mg/ml    ______________________________________    Milli-Q Water       0.0002    Ethanol             0.051    Benzyl alcohol      1.674    Propylene glycol    0.281    PEG 300             0.706    Dimethylisosorbide  0.928    Dimethylacetamide   5.000    N-Methyl-2-pyrrolidinone (NMP)                        >15.000 (range 15-20)    ______________________________________

NMP is inert with respect to undesirable chemical reactions with HLCDand is therefore a highly useful excipient to create solutions of HLCDin the lactone form. Further utility of NMP in the present invention isthe discovery that NMP allows the introduction of additional excipientswhich further improve the overall utility of the HLCD dissolved in theNMP solution in a manner which are of additional benefit for parenteralor oral administration to human patients with cancer. The HLCD solutionor suspension is prepared by mixing the desired components with NMP andadding a pharmaceutically acceptable acid to adjust the pH to 3-5.

REFERENCES

The following references may facilitate understanding or practice ofcertain aspects of the present invention. Inclusion of a reference inthis list is not intended to and does not constitute an admission thatthe reference represents prior art with respect to the presentinvention.

U.S. Patent

    ______________________________________    4,545,880      10/85      Miyasaka et al.    4,473,692      9/84       Miyasaka et al.    4,778,891      10/88      Tagawa et al.    5,061,800      10/91      Miyasaka et al.    4,772,589      9/88       Kaplan et al.    5,034,397      7/91       Kaplan et al.    ______________________________________

World Patent

    ______________________________________    9,014,094       11/90      Hui et al.    ______________________________________

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The foregoing description has been directed to particular embodiments ofthe invention in accordance with requirements of the Patent Statutes forthe purposes of illustration and explanation. It will be apparent,however, to those skilled in this art, that many modifications, changesand variations in the claimed antitumor compositions, solutions, methodsof administration of the antitumor compositions set forth will bepossible without departing from the scope and spirit of the claimedinvention. It is intended that the following claims be interpreted toembrace all such modifications and changes.

What is claimed is:
 1. A pharmaceutical formulation comprising an A-ringsubstituted camptothecin derivative having a water solubility of 5micrograms or less than 5 micrograms per milliliter, said A-ringsubstituted camptothecin dissolved or suspended inN-methyl-2-pyrrolidinone, said pharmaceutical formulation being suitablefor oral or parenteral administration to a patient.
 2. Thepharmaceutical formulation of claim 1, and further comprising apharmaceutically acceptable acid.
 3. The pharmaceutical formulation ofclaim 2, and further comprising a polyethylene glycol.
 4. Thepharmaceutical formulation of claim 3 wherein said polyethylene glycolis selected from the group consisting of PEG-300 and PEG-400.
 5. Thepharmaceutical formulation of claim 2, and further comprising anon-ionic surfactant.
 6. The pharmaceutical formulation of claim 5wherein said non-ionic surfactant is polysorbate-80.
 7. Thepharmaceutical formulation of claim 2, and further comprising a loweralcohol or benzyl alcohol.
 8. The pharmaceutical formulation of claim 7wherein said lower alcohol is ethanol.
 9. The pharmaceutical formulationof claim 1, and further comprising a pharmaceutically acceptable acid, alower alcohol, a polyethylene glycol, and a non-ionic surfactant. 10.The pharmaceutical formulation of claim 9 wherein said acid is citricacid, wherein said lower alcohol is ethanol, wherein said polyethyleneglycol is selected from the group consisting of PEG-300 and PEG-400, andwherein said surfactant is polysorbate-80.
 11. A pharmaceuticalformulation comprising an A-ring substituted camptothecin having a watersolubility of 5 micrograms or less than 5 micrograms per milliliter,wherein for each part by weight of substituted camptothecin saidsolution or suspension has 25 to 10,000 parts N-methyl-2-pyrrolidinone,100 to 5,000 parts of a pharmaceutically acceptable acid, 1,000 to10,000 parts by weight of polyethylene glycol, and 1,000 to 5,000 partsof a lower alcohol or benzyl alcohol.
 12. A pharmaceutical formulationcomprising a solution or suspension of an A-ring substitutedcamptothecin derivative having a water solubility of 5 micrograms orless than 5 micrograms per milliliter, wherein for each part by weightof the substituted camptothecin derivative said solution or suspensionhas 1,000 to 10,000 parts N-methyl-2-pyrrolidinone, 100 to 5,000 partsof a pharmaceutically acceptable acid, 1,000 to 5,000 parts of a loweralcohol, and 1,000 to 10,000 parts of a non-ionic surfactant.
 13. Thepharmaceutical formulation of claim 12 wherein said acid is citric acid,wherein said alcohol is ethanol, and wherein said non-ionic surfactantis polysorbate-80.